Many existing anti-cancer chemotherapeutics are non-specific, in that they typically damage or kill normal cells as well as malignant cells. Research in oncology is increasingly focused on targeted therapies, in which a therapeutic compound interacts with a specific molecule to interfere with a particular molecular pathway. Tumors in different individuals, even when found at the same anatomic location, can differ in their molecular signalling pathways. Determining which molecules and pathways are affected by a therapeutic compound provides techniques to select patients suitable for treatment with that therapeutic, as well as provides methods of monitoring therapy in order to identify patients whose tumors are responding to the particular therapy in use.
ErbB Receptors
The ErbB family of type I receptor tyrosine kinases includes ErbB1 (also known as the epidermal growth factor receptor (EGFR or HER1)), ErbB2 (also known as Her2), ErbB3, and ErbB4. These receptor tyrosine kinases are widely expressed in epithelial, mesenchymal, and neuronal tissues where they play a role in regulating cell proliferation, survival, and differentiation (Sibilia and Wagner, Science, 269: 234 (1995); Threadgill et al., Science, 269: 230 (1995)). Increased expression of wild-type ErbB2 or EGFR, or expression of constitutively activated receptor mutants, transforms cells in vitro (Di Fiore et al., 1987; DiMarco et al, Oncogene, 4: 831 (1989); Hudziak et al., Proc. Natl. Acad. Sci. USA., 84:7159 (1987); Qian et al., Oncogene, 10:211 (1995)). Increased expression of ErbB2 or EGFR has been correlated with a poorer clinical outcome in some breast cancers and a variety of other malignancies (Slamon et al., Science, 235: 177 (1987); Slamon et al., Science, 244:707 (1989); Bacus et al, Am. J. Clin. Path., 102:S13 (1994)).
A family of peptide ligands binds to and activates ErbB receptor signaling, and includes epidermal growth factor (EGF) and transforming growth factor α (TGFalpha), each of which binds to EGFR (Reise and Stern, Bioessays, 20:41 (1998); Salomon et al., Crit. Rev. Oncol. Hematol., 19: 183 (1995)). Ligand-receptor interactions are selective in that epidermal growth factor (EGF) and transforming growth factor alpha (TGFalpha) bind EGFR while heregulin binds ErbB3 and ErbB4. Ligand binding induces ErbB receptor phosphorylation (activation) with subsequent formation of homo- and heterodimers. ErbB2 is the preferred heterodimeric partner for EGFR, ErbB3, and ErbB4 (Graus-Porta et al., EMBO J., 16:1647 (1997); Tzahar et al., Mol. Cell. Biol., 16: 5276 (1996)). A number of soluble ligands have been identified for EGFR, ErbB3, and ErbB4, but none have been identified for ErbB2, which seems to be transactivated following heterodimerization (Ullrich and Schlessinger, Cell, 61: 203 (1990); Wada et al., Cell, 61: 1339 (1990); Karunagaran et al., EMBO J., 15:254 (1996); Stern and Kamps, EMBO J., 7: 995 (1988)).
ErbB1 and ErbB2 contain multiple tyrosine phosphorylation sites, and autophosphorylation of specific tyrosine residues within the highly conserved catalytic kinase domains of ErbB1 and ErbB2 establishes binding sites for Src-homology 2 (SH2) and phosphotyrosine-binding-domain containing proteins linking ErbB receptors to downstream cell proliferation (mitogen-activated protein kinase or MAPK; also known as Erk1/2) and survival (phosphatidylinositol-3-kinase or PI3K) pathways. Hackel et al: Curr Opin Cell Biol 11:184 (1999); Tzahar et al, Mol Cell Biol 16:5276 (1996); Lange et al. J Biol Chem 273:31308 (1998); Bacus et al., Oncogene 21:3532 (2002). Therapeutic modalities that target ErbB receptors and inhibit tyrosine kinase phosphorylation have been developed.
Therapeutics and ErbB2
Trastuzumab (Herceptin™), a humanized anti-ErbB2 monoclonal antibody has been approved for the treatment of breast cancers that either overexpress ErbB2, or that demonstrate ErbB2 gene amplification (Cobleigh et al, J. Clin. Oncol., 17:2639 (1999)). Trastuzumab binds to the extracellular domain of the ErbB2 receptor, and has been reported to exert its antitumor effects through several mechanisms. See e.g., Sliwkowski et al., Semin. Oncol. 26(Suppl 12):60 (1999).
Gefitinib is a small molecule that targets and inhibits phosphorylation of EGFR (ErbB1). Gefitinib is approved for third-line treatment of non-small cell lung cancer.
Because heterodimers of ErbB2 and EGFR can elicit potent mitogenic signals, interrupting both ErbB2 and EGFR simultaneously is a potential therapeutic strategy (Earp et al., Breast Cancer Res. Treat., 35:115 (1995)). Small molecule, dual EGFR-ErbB2 tyrosine kinase inhibitors have been identified and their pre-clinical anti-tumor activities reported (Fry et al., Proc. Natl. Acad. Sci. USA., 95:12022 (1998); Cockerill et al., Bioorganic Med. Chem. Letts., 11:1401 (2001); Rusnak et al., Cancer Res., 61:7196 (2001); Rusnak et al., Mol. Cancer Therap., 1:85 (2001)).
GW572016 (lapatinib) is a potent reversible, dual inhibitor of the tyrosine kinase domains of both EGFR and ErbB2, with IC50 values against purified EGFR and ErbB2 of 10.2 and 9.8 nM, respectively (Rusnak et al., Mol. Cancer. Therap., 1:85 (2001)). Recent reports have demonstrated that lapatinib inhibits EGFR and ErbB2 autophosphorylation in tumor cell lines that overexpress these receptors (Rusnak et al., Mol. Cancer. Therap., 1:85 (2001)), an effect that was primarily associated with tumor cell growth arrest. The chemical name of lapatinib (GW572016) is N-{3-chloro-4-[(3-fluorobenzyl)oxy]phenyl}-6-[5-({[2-methylsulfonyl)ethyl]amino}methyl)-2-furyl]-4-quinazolinamine (WO 99 35146, Carter et al.); a ditosylate form is disclosed in WO 02 02552 (McClure et al); methods of treating cancer are disclosed in WO 02/056912, and PCT/US03/10747.
It would be useful to identify biological molecules (biomarkers) that can be assessed, prior to therapy with a particular targeted therapy, to predict whether a tumor is likely to respond to that therapy. By screening subjects prior to therapy, those unlikely to respond to a given therapy can be treated with an alternate therapy. Similarly, it would be useful to identify biological molecules (biomarkers) that can be assessed during treatment with a particular targeted therapy, to indicate whether a tumor is responding to that therapy. By assessing such indicative biomarkers during therapy, resistance or non-response to a given therapy can be identified and an alternate therapy provided.